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Acinetobacter baumannii is one of the most notorious nosocomial pathogens with high mortality rates. Recently, egg yolk antibody (IgY), has been considered as a promising biomolecule against pneumonia caused by this bacterium. Loop 3 of outer membrane protein 34 (Omp34) was predicted as a highly exposed immunogenic peptide. However, its immunogenicity remains to be experimentally elucidated. In the current study, a construct composed of 5 copies of loop3 of Omp34 labeled as Omp34L3X5 was designed. This construct as well as the recombinant Omp34 were expressed, purified, and injected into laying hens to raise specific antibodies. The specific IgYs were extracted from hyperimmune egg yolks. The Omp34L3X5 and whole cells (WC) of A. baumannii served as antigens in indirect ELISA to assess the purified IgYs reactivity. These antibodies were administered to neutropenic mice 1 h before the challenge with 10 × LD

Type VI Secretion System (T6SS) contributes to both virulence and antimicrobial resistance in Acinetobacter baumannii. Valine-glycine repeat protein G (VgrG) is the core component of T6SS that exists in many bacterial pathogens that have emerged as a potent mediator of pathogenicity in A. baumannii. Two conserved sequences of vgrG 1263-2295 and vgrG1263-1608 were identified antigenic in various strains of Acinetobacter baumannii. The vgrg1263-1608 sequence was implanted in the Loopless C lobe (LCL) from N. meningitidis for surface display and exposure to functional epitopes. The VgrG and LCL-VgrG were expressed and purified. Groups of BALB/c mice were immunized with these proteins and challenged with A. baumannii. Specific IgG titers, whole-cell ELISA, animal survival rates in active and passive immunizations, the bacterial burden in mice tissues, and cytotoxicity of the proteins were determined. The specific IgG suppressed bacterial burdens in the organs, and increased survival rates were noted in the immunized mice. LCL-VgrG immunization provided better protection against A. baumannii infection than the VgrG immunization. The conserved region of VgrG is probably a safe immunogen to effective vaccine development or an antiserum to control A. baumannii infections.

The recent pandemic caused by a novel coronavirus known as SARS-CoV-2 has caught the international community by surprise. There is still no effective vaccine or treatment option against this virus. In this perspective, we discussed the potential protective and therapeutic effects of chitosan, as an FDA-approved biomolecule, against COVID-19 and influenza viruses.

Acinetobacter baumannii is a serious health threat with a high mortality rate. We have already reported prophylactic effects of IgYs raised against OmpA and Omp34 as well as against inactivated whole-cell (IWC) of A. baumannii in a murine pneumonia model. However, the infection was exacerbated in the mice group that received IgYs raised against the combination of OmpA and Omp34. The current study was conducted to propose reasons for the observed antibody-dependent enhancement (ADE) in addition to the therapeutic effect of specific IgYs in the murine pneumonia model. This phenomenon was hypothetically attributed to topologically inaccessible similar epitopes of OmpA and Omp34 sharing similarity with peptides of mice proteins. In silico analyses revealed that some inaccessible peptides of OmpA shared similarity with peptides of Omp34 and Mus musculus. Specific anti-OmpA and anti-Omp34 IgYs cross-reacted with Omp34 and OmpA respectively. Specific IgYs showed different protectivity against A. baumannii AbI101 in the murine pneumonia model. IgYs triggered against OmpA or IWC of A. baumannii were the most protective antibodies. IgY triggered against Omp34 is ranked next after those against OmpA. The lowest protection was observed in mice received IgYs raised against the combination of rOmpA and rOmp34. In conclusion, specific IgYs against OmpA, Omp34, and IWC of A. baumannii could serve as novel biotherapeutics against A. baumannii pneumonia.

Epsilon toxin (ETX) is one of four lethal toxins of Clostridium perfringens produced by types B and D of the pathogen. This pore-forming toxin is one of the most potent bioterrorism agents with economic importance. Although an effective vaccine and an equine antitoxin are available to protect livestock against ETX, no approved vaccine or antitoxin is available for humans. In the current study, an integrative, simple, fast, and reliable approach is availed to design a safe and minimized construct based on preexisting experimental linear B cell epitopes. This guideline is designed based on the surface accessibility, flexibility, hydrophilicity, content of beta-turn structure, and antigenicity of potential epitopes. Experimental linear B-cell epitopes were analyzed with respect to antigenicity. Two antigenic regions were introduced based on the distribution of epitopes. Moreover, three constructs were designed based on top ranking epitopes. These constructs were evaluated for their antigenicity, surface accessibility, flexibility, hydrophilicity, and beta-turn content and were compared to the ETX sequence. The 114-aa construct with an antigenicity score of 0.8592 was the most stable, antigenic, hydrophilic, flexible, and surface accessible antigen within the ETX sequence, the selected regions, and the designed constructs. Although the performed in silico analyses revealed that the designed construct could serve as a safe antigen triggering highly reactive and neutralizing anti-ETX antibodies, it should be verified by experimental assays in future studies.

Background: Cholera triggered by Vibrio cholerae remains the main reason for morbidity and mortality all over the world. In addition, salmonellosis is regarded as an infectious disease that makes it essential for the identification and detection of Salmonella. With a beta-barrel structure consisting of eight non-parallel beta strands, OmpW family is widely distributed among gram-negative bacteria. Moreover, OmpW isolated from S. typhimurium and Vibrio cholerae can be used in vaccine design. Methods: Topology prediction was determined. T-cell and B-cell epitopes were selected from exposed areas, and sequence conservancy was evaluated. The remaining loops and inaccessible residues were removed to prepare OmpW-1. High antigenicity peptides were detected to replace inappropriate residues to obtain OmpW-2. Physicochemical properties were assessed, and antigenicity, hydrophobicity, flexibility, and accessibility were compared to the native Omp-W structure. Low score areas were removed from the designed structure for preparing the OmpW-3. To construct OmpW-4, TTFrC was used as T-CD4+ cell-stimulating factor and CTB as adjuvant to the end of the C-terminal of this sequence, which can increase the antigenicity and sequence density. The sequences were re-analyzed to delete the unfavorable residues. Besides, the solubility of the mature OmpW and the designed structure were predicted while overexpressed in E. coli. Results: The designed vaccine is a stable protein which has immune cells recognizing epitopes and is considered as an antigen. The construct can be overexpressed in a E. coli. Conclusion: The multi-epitope vaccine is a suitable stimulator for immune system and would be a candidate for experimental research. Recent patents describing numerous inventions related to the clinical facets of vaccine peptide against human infectious disease.

The interaction between PD-L1 on tumor cells and the programmed death 1 (PD1) on immune cells helps them to escape the immune system elimination. Therefore, developing therapeutic agents to block this interaction has garnered a lot of attention as a therapeutic approach. In the present study, we have tried to screen for an inhibitory compound to inhibit the interaction between the PD1/PD-L1 molecules.In this regard, the structure of PD-L1 and its inhibitor were prepared and employed to generate an e-Pharmacophore model. A library of approved compounds was prepared and toxicity analysis using Absorption, Distribution, Metabolism, Excretion, and Toxicity (ADMET) predictor was performed. The built e-Pharmacophore model was validated and used to screen the prepared compound library. Ligand docking and binding energy calculation were performed on the screened ligands.A seven-feature e-Pharmacophore model was generated using the PD-L1 complex. All of the compounds within the library passed the ADMET criteria. Performing the virtual screening, only 79 compounds have survived the criteria to fit four pharmacophoric features. The compound with the highest binding energy was the liothyronine (T3).The ability of T3 in PD1/PD-L1 checkpoint blockade along with its potential in T4 reduction could be a desirable combination in cancer treatment. These abilities of T3 could be used to restore the ability of the immune system to eliminate tumor cells.

Acinetobacter baumannii is one of the most problematic pathogens in clinical settings. Emerging of its antibiotic-resistant strains persuade researchers to find alternative treatment options such as immunization against the notorious nosocomial pathogen. Oma87 has been introduced as an immunogenic outer membrane protein via reverse vaccinology. However, protectivity of A. baumannii Oma87 is not well known. The current research undertakes a study on the immunogenicity of recombinant Oma87 in a murine model. Some physico-chemical properties were assessed via in silico analyses. The corresponding gene was amplified and cloned into pET28a plasmid. The recombinant protein was purified and then was administered to immunize mice. Sera obtained from the immunized mice were assessed with respect to the triggered antibodies. Challenges were performed on actively or passively immunized mice. In silico analyses revealed that this protein is the same as BamA. A high titer of specific antibody was raised against rOma87 even after the first injection. The specific antibody recognized the whole cell of A. baumannii. Both active and passive immunizations confer 100 and 50% protection, respectively against ~ 2 × lethal dose (LD) of A. baumannii in the murine sepsis model. Although none of mice received ~ 5 × LD of A. baumannii survived in passive immunization, 25% of mice challenged with ~ 7 × LD of the bacteria survived and the dead mice exhibited a delayed death. Based on these results, Oma87 is the same as BamA which could be considered as a promising vaccine candidate against A. baumannii in the sepsis model. Electronic supplementary material The online version of this article (10.1007/s10989-020-10056-0) contains supplementary material, which is available to authorized users.

Membrane proteins play important functions, such as cellular communication and transferring materials in the cell. Many membrane proteins are involved in human diseases. However, little information is available on their structure, stability and folding. To study their structure, membrane proteins should primarily be produces in large amounts. However, production of these proteins is limited by various technical issues. Developing novel strategies to circumvent these issues seems to be highly important to proceed in membrane protein science. In this review, we have summarized the recent findings which can promote membrane protein expression and purification.

The complexity of treating Acinetobacter baumannii infections with the newly developed resistant strains has led researchers to confront this pathogen by developing vaccines. In this study, we used two important virulence factors of A. baumannii to elicit immunity against the A. baumannii. The immunogenic loops were from Baumannii acinetobactin utilization A (BauA) and 34kD outer membrane protein (Omp34). C-lobe derivative of the TbpB surface lipoprotein was used to display the superficial epitopes of the TbpA receptor protein of Neisseria meningitidis. The resulting loopless C-lobe (LCL) with implanted nucleotide sequences of the immunogenic loops from BauA and Omp34 was used as a hybrid antigen. The hybrid antigens were expressed in the E. coli and were used to immunize mice. The mice were challenged with a clinical isolate of A. baumannii (ABI022). Immunization with the hybrid antigens of the BauA loop 7 (BauAL7P3), Omp34 loop 3 Omp34L3P1, and the combination of both loops (BauAL7P3Omp34L3P1) brought about 42.86%, 42.86%, and 71.43% protection against A. baumannii infection. Histopathological findings in the immunized mice showed bronchioles clear from inflammatory cells and normal texture of the spleen and liver. The findings support the use of a multivalent vaccine to induce broadly reactive antibody responses against heterologous A. baumannii strains.

The structural consequences of ongoing mutations on the SARS-CoV-2 spike-protein remains to be fully elucidated. These mutations could change the binding affinity between the virus and its target cell. Moreover, obtaining new mutations would also change the therapeutic efficacy of the designed drug candidates. To evaluate these consequences, 3D structure of a mutant spike protein was predicted and checked for stability, cavity sites, and residue depth. The docking analyses were performed between the 3D model of the mutated spike protein and the ACE2 protein and an engineered therapeutic ACE2 against COVID-19. The obtained results revealed that the N501Y substitution has altered the interaction orientation, augmented the number of interface bonds, and increased the affinity against the ACE2. On the other hand, the P681H mutation contributed to the increased cavity size and relatively higher residue depth. The binding affinity between the engineered therapeutic ACE2 and the mutant spike was significantly higher with a distinguished binding orientation. It could be concluded that the mutant spike protein increased the affinity, preserved the location, changed the orientation, and altered the interface amino acids of its interaction with both the ACE2 and its therapeutic engineered version. The obtained results corroborate the more aggressive nature of mutated SARS-CoV-2 due to their higher binding affinity. Moreover, designed ACe2-baased therapeutics would be still highly effective against covid-19, which could be the result of conserved nature of cellular ACE2. Supplementary Information The online version contains supplementary material available at 10.1007/s10989-021-10346-1.

Acinetobacter baumannii causes severe nosocomial infections and is a difficult-to-treat pathogen due to the development of multidrug-resistant (MDR) strains. Vaccines and antibody therapy represent alternative promising strategies for the control of infections caused by A. baumannii or its MDR strains. OmpA and BauA have been assigned as protective antigens. However, the efficacy of the combination of these antigens is yet to be investigated. In this study, we targeted two critical antigens of A. baumannii (BauA and OmpA) to enhance immunoprotecting against A. baumannii.The recombinant BauA and OmpA were expressed and purified. The purified proteins were administered to BALB/c mice alone and in combination. Immune sera were assessed against BauA, OmpA and two constructs harboring immunogenic loops of these antigen. The mice were then challenged with a clinical isolate of A. baumannii. Indirect ELISA confirmed significant antibody rise to the antigens. The immunogenic loops were detected in the hybrid construct. The specific sera detected OmpA, BauA and constructs harboring immunogenic loops of these antigen with different affinities. A significant decrease in the bacterial loads was noted in the spleen, liver, and lungs of the immunized mice groups. However, the group received combination of BauA and OmpA showed lower bacterial burden in the spleen and liver.Combination of BauA and OmpA enhances immunoprotection against A. baumannii infections.

Acinetobacter baumannii is a common causative agent of nosocomial infections, with a mortality rate of 43% in infected patients. Due to the emergence of multidrug-resistant (MDR) strains, vaccine development has become necessary. Since the 34 kDa outer membrane protein Omp34 has been identified as a potential vaccine target, we implemented a hybrid antigen approach to target its extracellular loops. Using bioinformatic and structural analyses, we selected Loop 3 from Omp34 and displayed it on the loopless C-lobe (LCL) of TbpB of Neisseria meningitidis. The hybrid antigen and the LCL were produced and used to immunize mice for passive and active immunization and challenge experiments in which the reactivity of the sera was assessed by ELISAs, the bacterial load in the tissues measured and the survival of immunized mice compared. LCL was ineffective in immunization against A. baumannii thus the resulting immunity was due to the presence of Omp34 loop 3. It resulted in increased survival and a reduced bacterial load in the tissues compared to the control groups. The findings indicate that the immunogenicity of Omp34 loops can induce protection against A. baumannii infection, and it could probably be used as a vaccine candidate to control the pathogenesis of A. baumannii.

Acinetobacter baumannii, an opportunistic extracellular pathogen is one of the major causes of nosocomial infections. Omp34, also known as Omp33-36, is a bacterial porin protein involved in the virulence and fitness of this pathogen by adhesion to the host cell. This antigen nominated as an appropriate candidate for immunization against A. baumannii. In this study, the expression of the recombinant Omp34 (rOmp34) was carried out in E. coli BL21 (DE3). The immunogenicity of the rOmp34 in A. baumannii was studied in a murine sepsis model. Antibody response in mice injected with the recombinant protein was assessed using indirect ELISA. Bactericidal activity of rOmp34-immunized mice sera (1:10 dilution) against A. baumannii ATCC 19606 after 0, 1, 2, 4, and 8 h of incubation at 37 °C was assessed. In addition to survival rate, load of bacteria in liver and spleen of the infected mice were evaluated. A high titer of specific antibody equivalent to optical density of 1.54 ± 0.06 against rOmp34 was elicited in the immunized mice sera. Viability of the A. baumannii incubated 8 h with immunized mice sera was 64%. Homogenized liver and spleen samples of the control mice challenged with A. baumannii were loaded with 8 × 103 and 9 × 103 CFU per gram tissue respectively 48 h post-challenge as against complete clearance of A. baumannii in the immunized group. The protective immunity was achieved by challenging the mice groups with 5 × LD50 of live A. baumannii. Omp34 can be nominated as an immunogen that can bring about protection against Acinetobacter baumannii.

Autoimmune diseases (ADs) could occur due to infectious diseases and vaccination programs. Since millions of people are expected to be infected with SARS-CoV-2 and vaccinated against it, autoimmune consequences seem inevitable. Therefore, we have investigated the whole proteome of the SARS-CoV-2 for its ability to trigger ADs. In this regard, the entire proteome of the SARS-CoV-2 was chopped into more than 48000 peptides. The produced peptides were searched against the entire human proteome to find shared peptides with similar experimentally confirmed T-cell and B-cell epitopes. The obtained peptides were checked for their ability to bind to HLA molecules. The possible population coverage was calculated for the most potent peptides. The obtained results indicated that the SARS-CoV-2 and human proteomes share 23 peptides originated from ORF1ab polyprotein, nonstructural protein NS7a, Surface glycoprotein, and Envelope protein of SARS-CoV-2. Among these peptides, 21 peptides had experimentally confirmed equivalent epitopes. Amongst, only nine peptides were predicted to bind to HLAs with known global allele frequency data, and three peptides were able to bind to experimentally confirmed HLAs of equivalent epitopes. Given the HLAs which have already been reported to be associated with ADs, the ESGLKTIL, RYPANSIV, NVAITRAK, and RRARSVAS were determined to be the most harmful peptides of the SARS-CoV-2 proteome. It would be expected that the COVID-19 pandemic and the vaccination against this pathogen could significantly increase the ADs incidences, especially in populations harboring HLA-B*08:01, HLA-A*024:02, HLA-A*11:01 and HLA-B*27:05. The Southeast Asia, East Asia, and Oceania are at higher risk of AD development.

Acinetobacter baumannii is an important pathogen responsible for nosocomial infections worldwide. Trimeric autotransporters, the obligate homotrimeric adhesins, are involved in adherence of bacterial cells to various surfaces. These sorts of adhesins were shown to be expressed by A. baumannii. Trimeric autotransporters are modular virulence factors, containing numerous domains and structural architectures. The better understanding of the sequence and structural features of virulence factors are crucial in designing new therapeutic strategies. In this regard, with the aid of reliable in silico tools and the concept of “inherence through homology”, some sequence and structural features of AtaA.baumannii were unveiled. Domain architectures such as the position of repetitive modules, and coiled-coils, along with the prediction of tertiary and quaternary structures, allows us to define some important landscapes of AtaA.baumannii virulence factor. In addition, through CLANS analysis of TAA sequences of Moraxellaceae family, it was concluded that Ata contains several conserved of structural blocks of TAAs. The protein is initiated with extended signal peptide region, a stalk of several head domains and a membrane anchoring region. The globular heads are connected to each other by neck mediators and coiled-coil regions. Several fibronectin and collagen binding sites were defined within the structures. Minimal and maximal frustrated contacts are distributed within the structure of Ata, which suggest both the flexibility and toughness. These are come together for creating an efficient adhesin which is able to bind and bend through multiple sites. The protein could be great target for designing new vaccines or anti-virulence drug.

Although the biology of adrenocorticotropic hormone (ACTH) protein has already been scrutinized, some functional aspects of its biology are yet to be elucidated in the context of immunological disorders. In this regard, virtual screening of a compound library was performed against the structure of Cytotoxic T-Lymphocyte Associated Protein-4 (CTLA-4) (assessed both spatially and energetically) to discover novel biological functions for ACTH. The results of virtual screening and the MD simulation demonstrated that DB01284 has high binding energy along with proper interaction orientation against CTLA-4 (FG loop) by a clamp like structure. The employed methodology was checked using confirmatory control analyses. Intriguingly, DB01284 belongs to Tetracosactide (already prescribed protein drug for clinical conditions) which is the N-terminal region of ACTH. This is the first study to reveal that ACTH protein binds to the same amino acids of CTLA-4 (FG-loop) as B7 and anti-CTLA-4 antibody binds. In light of this finding, the molecular mechanism of ACTH function in patients suffering from Cushing’s Syndrom and the immunological bases for ACTH therapy of multiple sclerosis (MS) patients could be further delineated. Moreover, this finding suggests that ACTH could also act to block CTLA-4 in the context of anticancer immune check point blockade.

Highlights • Designed ACE2 lacks enzymatic activity and bears higher thermostability and affinity towards RBD. • The Thr27Arg mutation was introduced to increase the binding affinity against RBD. • The Arg273Gln and Thr445Gly mutations were introduced to reduce the ACE2 enzymatic activity. • The Asp427Arg mutation was done to decrease the flexibility of the ACE2. • The Pro451Met and Gly448Trp mutations were introduced to increase thermostability., The interaction between the angiotensin-converting enzyme 2 (ACE2) and the receptor binding domain (RBD) of the spike protein from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) plays a pivotal role in virus entry into the host cells. Since recombinant ACE2 protein has been suggested as an anti-SARS-CoV-2 therapeutic agent, this study was conducted to design an ACE2 protein with more desirable properties. In this regard, the amino acids with central roles in enzymatic activity of the ACE2 were substituted. Moreover, saturation mutagenesis at the interaction interface between the ACE2 and RBD was performed to increase their interaction affinity. The best mutations to increase the structural and thermal stability of the ACE2 were also selected based on B factors and mutation effects. The obtained resulted revealed that the Arg273Gln and Thr445Gly mutation have drastically reduced the binding affinity of the angiotensin-II into the active site of ACE2. The Thr27Arg mutation was determined to be the most potent mutation to increase the binding affinity. The Asp427Arg mutation was done to decrease the flexibility of the region with high B factor. The Pro451Met mutation along with the Gly448Trp mutation was predicted to increase the thermodynamic stability and thermostability of the ACE2. The designed therapeutic ACE2 would have no enzymatic activity while it could bear stronger interaction with Spike glycoprotein of the SARS-CoV-2. Moreover, decreased in vivo enzymatic degradation would be anticipated due to increased thermostability. This engineered ACE2 could be exploited as a novel therapeutic agent against COVID-19 after necessary evaluations.

Bacillus Calmette–Guerin (BCG) is an established vaccine used for tuberculosis (TB) prevention; however, it is not effective in adults and is only protective for children’s extrapulmonary TB. Therefore, many efforts have focused on producing effective TB vaccines for adults. One promising approach is DNA vaccination because of its ability to elicit both humoral and cellular immune responses, low cost, rapid production, desirable stability, and efficient protection against several infectious diseases. Nanoparticle (NP)-based systems can also enhance the immunogenicity of DNA vaccines by protecting naked plasmid DNA from nucleases and improving its delivery to immune cells. In this review, we focus on the design and delivery of novel DNA-based vaccines against TB and summarize investigations on specific mycobacterial antigens and chimeric combinations in animal models.

Objective: The current study compared the clinical, laboratory and radiological findings between groups of smokers and non- smokers Iranian patients with COVID-19. Methods: This was a case-control study done on 120 patients with COVID-19 that were admitted to Baqiyatallah hospital, Tehran, Iran during March to May 2020. Our patients were categorized into two groups: smokers (40 patients) and non-smokers (80 age and sex matched controls). Demographic and clinical characteristics, laboratory findings, imaging manifestations, and outcomes were compared between two age groups. Results: Regarding the comorbidities, no significant difference was observed between the smoker and non-smoker patients. The distribution of COVID-19 symptoms was not significantly different between smokers and non-smokers, except for chest pain and weakness, which were significantly more common in smokers with COVID-19. A significantly lower white blood cell count and neutrophils in peripheral blood sample of smokers however, no difference was found concerning lymphocyte count. Moreover, the RDW of smokers was significantly lower. Regarding the findings on CT scan of COVID-19 patients, no significant difference was found between smoker and non-smoker COVID-19 patients. One (2.5%) of deaths occurred in the smoker and 3 (3.8%) occurred in the non- smoker groups.Conclusion: Current findings showed that the clinical picture of smoker and non-smoker COVID-19 patients does not differ significantly.

Cholera triggered by Vibrio cholerae remains the main reason for morbidity and mortality all over the world. In addition, salmonellosis is regarded as an infectious disease that makes it essential for the identification and detection of Salmonella. With a beta-barrel structure consisting of eight non-parallel beta strands, OmpW family is widely distributed among gram-negative bacteria. Moreover, OmpW isolated from S. typhimurium and Vibrio cholerae can be used in vaccine design.Topology prediction was determined. T-cell and B-cell epitopes were selected from exposed areas, and sequence conservancy was evaluated. The remaining loops and inaccessible residues were removed to prepare OmpW-1. High antigenicity peptides were detected to replace inappropriate residues to obtain OmpW-2. Physicochemical properties were assessed, and antigenicity, hydrophobicity, flexibility, and accessibility were compared to the native Omp-W structure. Low score areas were removed from the designed structure for preparing the OmpW-3. To construct OmpW-4, TTFrC was used as T-CD4+ cell-stimulating factor and CTB as adjuvant to the end of the C-terminal of this sequence, which can increase the antigenicity and sequence density. The sequences were re-analyzed to delete the unfavorable residues. Besides, the solubility of the mature OmpW and the designed structure were predicted while overexpressed in E. coli.The designed vaccine is a stable protein that has immune cells recognizing epitopes and is considered as an antigen. The construct can be overexpressed in an E. coli.The multi-epitope vaccine is a suitable stimulator for the immune system and would be a candidate for experimental research. Recent patents describe numerous inventions related to the clinical facets of vaccine peptide against human infectious disease.

Acinetobacter baumannii, one of the most life-threatening nosocomial drug-resistant pathogens, imposes high morbidity and mortality rates, thus highlighting immunization-based treatments or prevention measures. The selection of appropriate antigens can elicit protective immunity. The gene encoding a fimbrial protein introduced via reverse vaccinology was cloned, expressed and evaluated for immunogenicity in a murine model. Mice immunized with the recombinant protein were challenged with A. baumannii ATCC 19606. Adherence to A549 cell line of specific anti-sera treated A. baumannii was also assessed. Passive immunity was evaluated in a murine pneumonia model. Indirect ELISA showed a high specific antibody titre. Adherence of A. baumannii to A549 cell line decreased by 40% after incubation with 1:250 dilution of specific anti-sera. All the actively immunized mice survived. Bacterial load in the spleen and liver of the immunized mice was 3-fold lower than those of the control. The number of bacteria in the lungs of passively immunized mice was about 6-fold lower than the control mice. The fimbrial protein could be considered as a promising protective immunogen against A. baumannii.

Nucleic acid immunization has recently exhibited a great promise for immunotherapy of various diseases. However, it is now clear that powerful strategies are imminently needed to improve their efficiency. In this regard, whole bacteriophage particles have been described as efficient DNA vaccine delivery vehicles, capable of circumventing the limitations of naked DNA immunization. Moreover, phage particles could be engineered to display specific peptides on their surfaces. Given these inherent characteristics of phages, we have designed a novel hybrid phage-DNA immunization vector using both M13 and pAAV plasmid elements. Following the construction and in vitro confirmation of the designed vectors, they were used for comparative mice immunization, carrying the same DNA sequence. The results indicated the efficacy of the designed hybrid phage particles, to elicit higher humoral immunity, in comparison to conventional DNA-immunization vectors (pCI). In light of these findings, it could be concluded that using adeno-associated virus (AAV) expression cassette along with displaying TAT peptide on the surface of the phage particle could be deemed as an appealing strategy to enhance the DNA-immunization and vaccination efficacy.

Acinetobacter baumannii, an emerging nosocomial pathogen, causes multi-drug and pan-drug resistant infections. This phenomenon necessitates the development of new treatment strategies or vaccines against this pathogen. Iron acquisition systems are important factors for the virulence of pathogenic organisms. Antibodies against iron regulated outer membrane proteins (IROMPs) showed bactericidal and opsonizing activities against A. baumannii in vitro. Data obtained from proteomic studies could be valuable for vaccine design. Comparative proteomic analysis of total lysate and outer membrane fractions isolated from A. baumannii ATCC 19606 cells cultured under iron-rich and chelated conditions resulted in the identification of protein spots differentially produced. Need for bioinformatic tools is imperative for analysis of these data as well as to design novel proteins bearing desired properties. This study undertakes in silico identification of the most effective immunogenic target amongst these iron regulated proteins in A. baumannii which can be employed as a vaccine candidate. Here a screening was carried out on iron- regulated A. baumannii OMPs, based on hydrophilicity, flexibility, beta-turns, solubility and overall antigenic probability. 3D structure of the proteins was modeled. Linear and conformational B cell epitopes predicted and their densities were used for comparison of their immunogenicity. CarO was selected as an efficient immunogenic IROMP in A. baumannii which contributes to sidrophore mediated iron uptake.

After years of ongoing endeavors for HBV infection prognosis, diagnosis and treatment, it still remains a major health problem worldwide. About 400 million chronic carriers and an annual death rate as high as one million reflects the seriousness of the problem. Developing novel and more effective diagnostic strategies, using in silico approaches and their subsequent empirical verification, will be helpful in providing for blood supply safety, therapeutics efficacy and disease activity assessment. Exploiting various in silico tools a novel multiepitope detection construct was designed which was consisted of eight linked linear immunodominant HBV epitopes. The designed antigen was expressed in Escherichia coli as the host. The detection capability of the designed antigen was tested using Chemiluminescent immunoassay method. Chemiluminescent immunoassay on the expressed antigen revealed that the product may be a credible candidate for simultaneous detection of three main HBV antibodies. All three test samples in two concentrations indicated lower RLU/s in comparison to the positive control which was the direct consequence of HBV antibody detection by the designed antigen. In the present study, employing bioinformatics tools paved the way for rational design of multiepitope antigen in a more cost effective, intelligent and knowledge-based method. The obtained results could be construed as a primary proof of concept that the in silico predictions could be used as primary steps of the biological studies and their subsequent empirical conduction.

Immunotherapy has been suggested as a compelling alternative approach for conventional breast cancer treatment methods. Despite the paramount rolesof T cells in this approach, insufficient numbers of them in the combat against progressive tumor growth still remain to be dealt with. Super antigens are a class of antigens, capable of eliciting T cell proliferation response against desired antigens. Staphylococcal enterotoxin B (SEB) is categorized as a super antigen, its anti-tumor properties has been previously reported. However, to the best our knowledge, SEB has not been ever administered as a DNA construct. In the present study, we exploited bioinformatics tools to assess the immunoreactivity of a SEB-coding DNA construct that serves as a DNA vaccine for breast cancer therapy. Potential B and T (MHC class I and II binders) cell epitopes of the hypothetically expressed protein, along with its sub cellular localization were predicted. Moreover, probable glycosylation and phosphorylation sites within the protein sequence were determined. The gene sequence was optimized according to murine model codon bias and its mRNA stability was analyzed. Employing an integrative in silico approach, we revealed that apparently the construct could be efficiently expressed in mouse model. Moreover, the hypothetically expressed protein could act as an amenable adjuvant in cancer immunotherapy.

Epsilon toxin of the Clostridium perfringens garnered a lot of attention due to its potential for toxicity in humans, extreme potency for cytotoxicity in mice and lack of any approved therapeutics prescribed for human. However, the intricacies of the Epsilon toxin action mechanism are yet to be understood. In this regard, various in silico tools have been exploited to model and refine the 3D structure of the toxin and its two receptors. The receptor proteins were embedded into designed lipid membranes within an aqueous and ionized environment. Thereafter, the modeled structures subjected to series of consecutive molecular dynamics runs to achieve the most natural like coordination for each model. Ultimately, protein-protein interaction analyses were performed to understand the probable action mechanism. The obtained results successfully confirmed the accuracy of employed methods to achieve high quality models for the toxin and its receptors within their lipid bilayers. Molecular dynamics analyses lead the structures to a more native like coordination. Moreover, the results of previous empirical studies were confirmed, while new insights for action mechanisms including the detailed roles of Hepatitis A virus cellular receptor 1 (HAVCR1) and Myelin and lymphocyte protein (MAL) proteins were achieved. In light of previous and our observations, we suggested novel models which elucidated the existing interplay between potential players of Epsilon toxin action mechanism with detailed structural evidences. These models would pave the way to have more robust understanding of the Epsilon toxin biology, more precise vaccine construction and more successful drug (inhibitor) design.

Trimeric autotransporter (TAA), also known as type Vc secretion system, is expressed by many strains of Acinetobacter baumannii, an opportunistic pathogen, which is responsible for nosocomial infections worldwide. TAAs, are modular homotrimeric virulence factors, containing a signal peptide, complex stalk, and conserved membrane anchoring domain. The evolutionary mechanisms underlying the evolvement of these adhesins are not clear. Here, we showed that TAA genes were laterally acquired and underwent gene duplication and recombination. The heterogeneity of TAA nucleotide sequences, GC content, codon usage, and the probability of recombination and duplication events were assessed by MEGA7. Given the heterogeneity of sequences, we used all-against-all BLAST for clustering the TAAs. The pattern of distribution of TAAs are highly scattered; GC content and codon usage for these genes are variable. Multiple events of lateral gene transfer from the early history of Acinetobacter and the occurrence of gene duplication, gene loss, and recombination after acquiring the alien genes may explain the scattered pattern of distribution of TAAs. Additionally, this gene is not present in many clinical isolates of A. baumannii, thus is not a single virulence factor attributing to the infection. The advantage of harboring such genes might be adopting to different environments by developing the biofilm communities. We suggested that TAA genes were laterally acquired in the environmental context and incidentally provided some benefits at the infection site. Thus, coincidental evolution theory may be better suited for describing the evolution of TAA genes in A. baumannii genomes.

Background Infections caused by drug-resistant strains of Acinetobacter baumannii and Pseudomonas aeruginosa are now a global problem that requires the immediate development of new antimicrobial drugs. Combination therapy and using antimicrobial peptides are two strategies with high potential to solve this issue. By these strategies, this study aimed to determine the antimicrobial effect of Nisin and P10 antimicrobial peptides on extensively drug-resistant Acinetobacter baumannii and colistin-resistant Pseudomonas aeruginosa isolates, and investigate the most effective combination of an antimicrobial peptide with an antibiotic. Material and methods This study was performed on five resistant clinical isolates and one standard strain for each kind of bacterium. First, the minimum inhibitory concentrations of two antimicrobial peptides (Nisin and P10) and five common antibiotics for the treatment of Gram-negative bacteria (ceftazidime, tobramycin, ciprofloxacin, doripenem, and colistin) was determined using Scanner-Assisted Colorimetric MIC Method. Then, the combination effect of P10+Nisin, P10+antibiotics, Nisin + antibiotics was investigated using checkerboard method. Results The MIC value of Nisin and P10 against studied pathogens were 64–256 and 8–32 μg/ml, respectively. P10+Nisin combination showed synergistic effect against standard strains and additive effect against drug-resistant clinical isolates. It was also found that the combination effect of P10+ceftazidim, P10+doripenem, and Nisin + colistin was synergistic in most cases. Nisin + tobramycin combination showed synergistic effect in exposure to standard strains, while the synergy is strain-dependent against drug-resistant clinical isolates. Conclusion In conclusion, the synergism of Nisin + colistin and P10+ceftazidime/doripenem could be of great therapeutic value as antimicrobial drugs against infections caused by colistin-resistant P.aeruginosa and XDR A. baumannii.